This invention relates to antigens expressed by hematopoietic stem cells and progenitor cells and to methods of using such antigens, especially for cell separation and purification.
The high turnover of mammalian blood cells requires a supply of hematopoietic stem cells that are able to give rise to other blood cell lineages. The immediate progeny of the hematopoietic stem cell are called progenitor cells, and are capable of giving rise to various cell types within one or more lineages, i.e. the erythroid, myeloid and lymphoid lineages. The stem cell and progenitor cell populations constitute only a small percentage of the total number of cells in bone marrow, fetal liver, etc. These populations are of immense interest because of their ability to repopulate the hematopoietic system.
A number of methods have been described in the literature for the purification or enrichment of hematopoietic stem cell and progenitor cell populations. There is significant commercial interest in these methods because hematopoietic progenitors have a number of clinical uses. Progenitor cell transplantation is currently used in conjunction with chemotherapy and radiation for the treatment of leukemia, breast cancer and other tumors. Frequently, autologous transplants are used to avoid the danger of graft rejection, but there is an increased risk of disease reappearance, due to the presence of tumor cells in the engrafting cell population. Transplantation of a more purified source of progenitor cells is therefore preferable.
There is also interest in the use of hematopoietic progenitor cells as a vehicle for gene therapy. Although not yet proven in the clinic, the longevity of hematopoietic stem cells and the dissemination of their progeny in the vasculature are desirable characteristics. A number of vectors, including several retrovirus and adenovirus based constructs, that can transfect hematopoietic stem cells have been described.
Proteins and other cell surface markers found on hematopoietic stem cell and progenitor cell populations are of great interest, as they are useful in preparing reagents for identification, separation and isolation of these populations and in the further characterization of these important cells. Although some antigens are now known that can be used in the identification and separation (positive and negative) of stem cells, such as (for example) the CD 34 antigen, which is found on stem cells but not on mature blood cells, there is a continued need for development of other antigens, particularly one that can simplify the identification and separation of desirable classes and subclasses of cells, especially hematopoietic stem cells and progenitor cells.
U.S. Pat. No. 5,061,620 describes a substantially homogeneous human hematopoietic stem cell composition and the manner of obtaining such composition. Stromal cell-associated hematopoiesis is described by Paul et al. (1991) Blood 77:1723-1733. The phenotype of stem cells with rhodamine staining is discussed in Spangrude and Johnson (1990) P.N.A.S. 87:7433-7437. Cell surface antigen expression in hematopoiesis is discussed in Strauss et al. (1983) Blood 61:1222-1231 and Sieff et al. (1982) Blood 60:703-713. Descriptions of pluripotential hematopoietic cells are found in McNiece et al. (1989) Blood 74:609-612 and Moore et al. (1979) Blood Cells 5:297-311. Characterization of a human hematopoietic progenitor cell capable of forming blast cell-containing colonies in vitro is found in Gordon et al. (1987) J. Cell. Physiol. 130:150-156 and Brandt et al. (1988) J. Clin. Invest. 82:1017-1027. The use of progenitor cells in transplantation is discussed in To et al. in Progenitor Threshold in Transplantation (ISBN 1-880854 17-1) pp. 15-20. Utilities for the cell compositions obtained using the methods and compositions of the invention are described in these publications, among others.
The use of high-gradient magnetic separation for the isolation of human hematopoietic progenitor cells is described in Thomas and Landsdorp (1992) in Advances in Bone Marrow Purging pp.537-544; and Kato and Radbruch (1993) Cytometry 14:384-392. Other methods of magnetic selection for human hematopoietic progenitor cells are described in Bigas et al. (1992) in Advances in Bone Marrow Purging pp.545-551; Oku et al. (1992) in Advances in Bone Marrow Purging pp. 553-560; and Hardwick et al. (1992) in Advances in Bone Marrow Purging pp. 583-589. High gradient magnetic cell sorting is described in Miltenyi et al. (1990) Cytometry 11:231-238. Molday, U.S. Pat. No. 4,452,773 describes the preparation of magnetic iron-dextran microspheres and provides a summary describing the various means of preparation of particles suitable for attachment to biological materials.
Methods and compositions are provided for the enrichment and characterization of human hematopoietic progenitor and stem cells. An antigen has been identified, referred to here as the AC133 antigen, that is present on stem cells and on progenitor cells and that can be used for the identification and/or separation of these important cells from the vast majority of cells present in a biological (or other) source of hematopoietic cells. Novel antigen compositions and reagents that react with them, such as antibodies, are provided for use in the methods of the invention and for the further investigation of hematopoietic progenitor and stem cell biology. For example, hematopoietic cells can be obtained from various sources, including fetal and adult bone marrow, cytokine mobilized peripheral blood cells, and fetal liver, and can be separated using reagents and methods of the invention.